Generation of Small Fusion Genes Carrying Phleomycin Resistance andDrosophilaAlcohol Dehydrogenase Reporter Properties: Their Application in Retroviral Vectors

We have used theDrosophilaADH cDNA to engineer new fusion genes carrying both reporter activity and bleomycin/phleomycin resistance (Sh ble). Cassettes of ADH::Sh ble, Sh ble::ADH, or ADH::Sh ble::ADH with or without polyadenylation signals were constructed. Placed under the control of the strong CMV promoter, these constructs induced intense ADH substrate staining and phleomycin resistance, whatever the position of the ADH gene, in avian or mammalian cell lines. SNV-based nonreplicative retroviral vectors were constructed and introduced into the appropriate packaging cell line. Titers up to 10⁶ADH forming units/ml of viral supernatant were obtained except for the ADH::Sh ble::ADH construct, which reached 10⁵ADH forming units. These retroviral vectors were inoculated to the E3 chick embryo via the coelom. Three days later, cells from different organs were put in culture for 24 h and stained to detect ADH activity. A large number of positive cells were found in cultures from all organs. The new fusion genes described here are, to our knowledge, the smallest (1.1 kb) published to date that carry both reporter and drug resistance properties. These genes represent the basis of a new retroviral vector model with three distinct properties in two genetic units; their advantage is to reduce the size and increase the efficiency of the vector.     

Analysis of an optimal hidden Markov model for secondary structure prediction

Background  

Secondary structure prediction is a useful first step toward 3D structure prediction. A number of successful secondary structure prediction methods use neural networks, but unfortunately, neural networks are not intuitively interpretable. On the contrary, hidden Markov models are graphical interpretable models. Moreover, they have been successfully used in many bioinformatic applications. Because they offer a strong statistical background and allow model interpretation, we propose a method based on hidden Markov models.     

Comparison of Growth Plasticity in the Laboratory and Field, and Implications for for the Onset of Juvenile Development in Sofie, Chondrostoma toxostoma

The aim of the present study was to determine the effect of environmental conditions (controlled and natural) on the duration and sequence of developmental steps in sofie, Chondrostoma toxostoma, early ontogeny. Few previous studies on the early development of fishes have included relative growth and none have compared relative growth in the laboratory and the field. Such comparison is important to quantify the morphological development of different parts of fish during their early ontogeny, to determine potential variations in growth that may occur under laboratory conditions and to understand better the plastic nature of relative growth. Early development and relative growth of 23 characters were examined in specimens of sofie reared under both laboratory and natural conditions in tributaries of the River Garonne basin (France). The sofie is still present in this basin despite progressive localised extinction in the rivers of south western Europe over the last 30 years. Growth of field and laboratory embryos (in degree days, °days) was the same up to larva step 1 (9 mm SL), but thereafter was markedly slower in the laboratory than in the field. Ontogenetic rate in the field was twice that in the laboratory, suggesting a precocial (specialist) form under natural conditions and an altricial (generalist) form under laboratory conditions. Stabilisation of relative growth, i.e. end of the remodelling process (metamorphosis), occurred well after all larval characteristics (remnants of finfold, rapid allometric growth) had disappeared and all the juvenile structures had appeared (nasal septa, complete scale cover). In the field, this stabilisation occurred in specimens of approximately 50 mm SL, suggesting that metamorphosis ends and the juvenile period begins at the end of the sofie's first (0+) year of life.     

Early Development of the Sofie, Chondrostoma toxostoma

The developmental biology of embryos, larvae and juveniles of sofie, Chondrostoma toxostoma, reared from artificial insemination, was examined under controlled laboratory conditions, using both in-vivo and preserved specimens. The complete remodelling process (metamorphosis) was described and interpreted according to the theory of saltatory ontogeny, providing a model within which the highly complex ecological niche and behaviour (habitat use, diet, etc.) of the free embryos and larvae can be evaluated. The sofie ova were relatively opaque and the free embryos presented a small yolksac. Consequently, the free embryo steps were short and the free embryos emerged rapidly from the substratum. The circulatory system appeared early and was simple, which suggests that sofie free embryos require well oxygenated water. The remodelling of the mouth from a superior to inferior position characterised changes in the sofie's diet during early life history, which is typical of the genus. Differences in development relative to the nase, C. naus, are discussed.     

Occurrence of somaclonal variation among somatic embryo-derived white spruces (Picea glauca,Pinaceae)

Four different variegata phenotypes were identified among 2270 white spruce plants [Picea glauca (Moench.) Voss.] produced over a period of 2 yr from the time of induction of embryogenic tissues. The four variegated plants differed from each other in the extent and distribution of chlorophyll-deficient needles. Light microscopy showed that variegated leaves of a selected variant consisted of a chimeral mixture of green and white cells. Electron microscopy showed that cells in completely white needles had large nuclei with predominant euchromatin, lacked large cytoplasmic vacuoles, and harbored vacuolized plastids with aberrant morphologies. Various observations suggest that the recovered variegata phenotypes reflect some kind of genetic instability of either chloroplastic or nuclear genomes. To elucidate the genetic basis of these variegata phenotypes in white spruce, three out of the four variants were subjected to randomly amplified polymorphic DNA (RAPD) analysis. Out of more than 250 RAPD markers screened, only one correlated with white needles of variegated plants. The nucleotide sequence of this DNA fragment showed no homology with any known gene, but the amplified sequence appears most likely of nuclear origin.     

Enamel microstructure evolution in anthracotheres (Mammalia,Cetartiodactyla) and new insights on hippopotamoid phylogeny

Investigations on enamel microstructure provided new data for the debate on hippopotamid origin. Observations indicated a diversity of patterns relevant to phylogenetic inferences. Within Hippopotamoidea, the distribution of these patterns seems to be in favour of a hippopotamid origin within the Palaeogene African anthracotheres. Enamel microcharacters therefore prove to be particularly relevant for future phylogenetic analysis of the superfamily, and have implications for our understanding of ecological transitions within hippopotamoids at the end of the Miocene. Indeed, unlike equids or bovids, which developed grass feeding thanks to their hypsodont molars, hippopotamoids may have had another way to exploit this resource. The combination of inter‐row sheets, which appeared early in the evolutionary history of the group, and the increased thickness of radial enamel could have eased the consumption of highly abrasive graminoids. © 2014 The Linnean Society of London     

ETH 6022: an artificial enzyme? A comparison between enzymatic and chemical recognition for sensing ethanol

Routinely the quantification of ethanol in clinical chemistry and food chemistry is performed by enzymatic methods in addition to chemical methods. Liver and yeast alcohol:NAD⁺ oxidoreductase (ADH, EC. 1.1.1.1) and alcohol oxidase AOD (EC 1.1.3.13) are mainly applied for selective evaluations by different kinds of bioassay. Nevertheless, drawbacks of ethanol sensing make the evaluation of ethanol concentrations in alcoholic beverages by an optical sensor, in a concentration range between 0·7% and 40%, most attractive. The chemical recognition process is based on a lipophilic amide of trifluoroacetylaniline (ETH 6022) as a ligand for ethanol implanted in a solvent polymeric membrane. The interaction leads to a hypsochromic shift of the absorption band in the ultraviolet region (305 nm). The chemical reaction in the apolar environment of the optode membrane is reversible. The principle of this recognition process is compared to the enzymatic reaction of ADH. In both cases a highly electrophilic centre participates in the interaction between the hydroxy group of the alcohol and the ligand. The selectivity behaviour of the artificial recognition process may be tailored by the substituents of the aromatic trifluoroacetyl compound. The response characteristics of the optochemical sensors are compared.